Peptides for inducing apoptosis in tumor cells

ABSTRACT

The present invention relates to peptides which interact with IAPs. IAPs are highly expressed in tumor cells which fail to undergo apoptosis. By binding to IAPs, the peptides of the present invention release tumor cells from the apoptosis block and thus provide a new tool for effective cancer therapy.

The present invention relates to peptides that are suitable to induceapoptosis, the nucleic acids encoding said peptides and the use of thepeptides and/or nucleic acids in cancer therapy.

Cancer is the second major cause of death in Europe and NorthernAmerica. The effective cure of patients, though, is often difficultsince many tumor cells have developed a resistance to anti-cancer drugsused for chemotherapy. The described phenotype involves a variety ofstrategies that tumor cells use to evade the cytostatic effects ofanticancer drugs. Mechanisms for drug resistance include modificationsin detoxification and DNA repair pathways, changes in cellular sites ofdrug sequestration, decreases in drug-target affinity, synthesis ofspecific drug inhibitors within cells, and accelerated removal orsecretion of drugs. Cancer cells commonly fail to undergo so-called“programmed cell death” or “apoptosis”, a signaling process that plays akey role in preventing cell tissues from abnormal growth. Thus,apoptosis defects appear to be a major problem in cancer therapy as theyconfer resistance to many tumors against current treatment protocols,leading to tumor progression.

Apoptosis pathways involve diverse groups of molecules. One set ofmediators implicated in apoptosis are so-called caspases, cysteineproteases that cleave their substrates specifically at aspartateresidues. Caspases convey the apoptotic signal in a proteolytic cascade,with caspases cleaving and activating other caspases which subsequentlydegrade other cellular targets eventually resulting in cellularbreakdown. If one or more steps in this cascade is inhibited in tumorcells, these cells fail to accomplish apoptosis and, thus, continue togrow. Caspase activation itself can be triggered by external stimuli orby intracellular stress response via the mitochondria leading to therelease of mitochondrial proteins.

A failure in activating the caspase cascade is caused by the action ofso-called Inhibitors of Apoptosis Proteins (IAPs). IAPs bind to earlyactive caspases, thereby preventing the ongoing of the apoptosisprocess. They are expressed at high levels in many tumors and, byinhibition of caspases, contribute to the resistance of cancers againstapoptosis induction.

From the foregoing it becomes evident, that inhibition of LAP functionrepresents a major task for efficient cancer therapy. It has beenreported earlier that the mammalian mitochondrial protein Smac, whenreleased into the cytosol in the course of an apoptotic response, canbind to IAPs and, thus, promotes the proteolytic activation of caspasesresulting in apoptosis. Therefore, Smac or fragments thereof, could be apotential tool for the treatment of drug-resistant tumors. However, thetherapeutic use of naturally existing LAP-binding proteins like Smac canbear the disadvantage of evoking undesired side effects. In addition,binding of Smac to IAPs is restricted to a particular region on the LAPprotein, the so-called BIR region. The problem underlying the presentinvention is to find other molecules which are able to promote apoptosisin tumor cells by binding to IAPs, but which do not possess thedisadvantages of naturally existing LAP-binding partners.

Peptides that bind to IAPs, in particular to the BIR region of IAPs areknown to the person skilled in the art. WO-A 02/16402 disclosesaminoterminal Smac peptides (so-called AV peptoides) which bind to IAPsand abolish their inhibitory effect. The amino acid sequence of thesepeptides follows a distinct order, namely that the first amino acid isan alanin, the second amino acid is preferably a valin, isoleucin orleucin, and the third amino acid is preferably a prolin or an alanin.WO-A 02/26775 discloses synthetic tetrapeptides, which also bindspecifically to the BIR domain of IAPs. The disclosed amino acidsequence of the tetrapeptides follows the order of alanine, valin (orthreonin or isoleucin), prolin (or alanin), phenylalanine (or tyrosin orisoleucin or valin). The problem of the present invention is to findpeptides, that do inhibit the function of IAPs via a differentmechanism.

The present invention solves the described problem by providingrandomized peptides that bind to particular IAPs and, thus, can impedesaid IAPs from exerting their anti-apoptotic effect.

The object of the present invention is attained by a peptide, a fragmentor derivative thereof, comprising an amino acid sequence selected fromthe group of SEQ ID NOS: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14,15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32,33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50,51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68,69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86.(see TABLE 1).

In the context of the present invention, the term derivative or fragmentof peptides further includes peptides in which one or more amino acidsof the disclosed sequences can be substituted by one or more amino acidsdifferent from the original one(s), or peptides the amino acid sequenceof which is either extended, shortened, or both, on either theaminoterminal, or the carboxyterminal or both ends with respect to theoriginal proteins, provided that the binding properties of the peptidesremain unaffected. Preferably they differ from the original amino acidsequence as disclosed in the present invention in no more that 50%, morepreferably not more than 35% and most preferably not more than 10%.

In a preferred embodiment of the present invention, the term derivativeor fragment particularly refers to a peptide comprising an amino acidsequence selected from the group of SEQ ID NOs:87, 88, 89, 90, 91, 92,93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108,109, 110, 111, 112, 113, 114, 114, 115, 116, 117, 118, 119, 120, 121,122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132 (see TABLE 2).

More preferably, the peptide, a fragment or derivative thereof, of thepresent invention comprises an amino acid sequence selected from thegroup of SEQ ID NOs:90, 91, 98, 99, 111, 118, 123, 124, 127, 128, 132.These SEQ ID NOs directly resemble numbers 4, 5, 12, 13, 25, 32, 37, 38,41, 42, 46 in TABLE 2 (or peptide numbers 5, 7, 15, 16, 40, 54, 65, 69,75, 77, 90 according to the inventors' numbering system). Particularlypreferred is a peptide comprising the SEQ ID NO: 127 (corresponds to No.41 in TABLE 2, or, respectively, peptide number 75 according to theinventors numbering system).

In the context of the present invention, the term derivative or fragmentof peptides further includes peptides which are shortened at theircarboxyterminal end. The inventors have found that, for example, aminoacids alanin, glutamic acid, isoleucin, tyrosin, glutamic acid, serin(A-E-I-Y-E-S), which are the last six carboxyterminal amino acids ofmost peptides of SEQ ID NOs: 87-132, can be omitted, without losing thecharacteristic functional properties of the respective peptides.However, it is also possible that the peptides of the present inventionare carboxyterminally shortened by more than these six amino acid,provided that the binding and functional properties of the peptidesremain unaffected.

According to a preferred embodiment of the present invention, thepeptide of the present invention, the fragment or derivative thereof isnot:

-   -   (i) a peptide comprising AX₁, wherein X₁ is V, I, or L    -   (ii) a peptide comprising AX₁X₂, wherein X₁ is V, I, or L,        preferably V, and X₂ is P or A, preferably P.    -   (iii) a tetrapeptide    -   (iv) a tetrapeptide having the sequence X1-X2-X3-X4, wherein X1        is A, X2 is V, T or I, X3 is P or A, and X4 is F, Y, I or V.

The peptides of the present invention bind to IAPs. Preferably they bindto human LIPs selected from the group consisting of c-IAP1, c-IAP2,XIAP, NAIP, survivin and livin/ML-LIP, and/or a fragment or derivativeof any of the aforementioned IAPs. Livin/ML-IAP will be referred to aslivin hereinafter. More preferably, the peptides bind to sunrivin andlivin, and most preferably said peptides bind to livin.

It is important to note that the peptides of the present invention donot interact with livin via their amino terminal end, as it has beendescribed in the art for other LIP-binding proteins like for exampleSmac/DIABLO. The amino terminal end of the peptides of the presentinvention is not freely accessible for any possible interaction with anIAP, since it is fused (for experimental purposes) to activation domainof the so-called GAL4 transcription activator (see EXAMPLES). Thus, itis obvious to the skilled artisan that the peptides of the presentinvention differ from those described in the art.

The peptides of the present invention can be identified by methods wellknown to the person skilled in the art. In general, the identificationis achieved by contacting a peptide library with the desired interactionpartner, e.g. livin, and selecting those which successfully bind. Amongthe known methods are screening of peptides libraries by e.g. phagedisplay, ribosome display, mRNA display and yeast and/or mammaliantwo-hybrid systems. The preferred method to identify the peptides of thepresent invention is the two-hybrid approach. Particularly preferred isthe use of the two-hybrid system to perform a so-called “peptide aptamerscreening” (see EXAMPLES).

For peptide aptamer screening, the peptides of the library canoptionally be anchored at both ends within a so-called scaffold protein,leading to a more preferred conformational stage of the peptides. Knownscaffold proteins comprise e.g. E. coli thioredoxin A (trxA),staphylococcal nuclease, protease inhibitor eglin, Tendamistat fromStreptomyces tendea, cellular transcription factor Sp1, and greenfluorescent protein GFP. The preferred scaffold protein of the presentinvention is trxA.

Optionally, the peptides can be linked to a second moiety to create aso-called fusion protein. The fusion partner can be a carrier, which ispreferably a protein, a fragment or derivative thereof, the attachmentof which to any of the peptides, fragment or derivative thereof enablesthe penetration of the peptides through the cell membrane into the cell.Appropriate carriers, in particular proteins, are known to the personskilled in the art and include TAT, influenza virus hemagglutinin, theVP22 protein from herpes simplex virus, Antennapedia, fibroblast growthfactor, Galparan (transportan), poly-arginine, Pep-1. Other carriersknown to a person skilled in the art which do not belong to proteins,but mediate the internalization of molecules into cells include lipidsand cationic lipids.

When a protein is used as a carrier, the term derivative or fragment ofa protein refers to peptides in which one or more aminoacids can besubstituted by other aminoacids different from the original one(s), orpeptides the aminoacid sequence of which is either extended, shortened,or both, on either the aminoterminal, or the carboxyterminal or bothends, with respect to the original one(s), provided that the function asa carrier for the cellular uptake of the peptides remains unaffected.The above definition relates to TAT, influenza virus hemagglutinin, theVP22 protein from herpes simplex virus, Antennapedia, fibroblast growthfactor, Galparan (transportan), poly-arginine and Pep-1.

In a preferred embodiment of the present invention, the carrier thepeptide is fused to is poly-arginine, particularly R7, R9 or R11(peptides with 7, 9 or 11 arginine residues).

The linkage of the peptides to the carrier can occur by any chemicalinteraction known to the person skilled in the art, like chemicaladsorption, dipole-dipole or the like. Preferably, the carrier is linkedto the peptides by a chemical bond, in particular a covalent bond, incase the carrier is a protein. This bond must be such that it remainsunaffected before and while penetrating the cell membrane and, ifnecessary for the interaction of the peptides with IAPs, can be cleaved.In general, the peptide/carrier entity can interact with IAPs to thenecessary extent, a cleavage being not necessary.

It is also possible that the carrier, also known to the skilled artisanas protein transduction domain (PTD), is linked to the peptide of thepresent invention via disulfide bonds instead of covalent linkage. Thisbears the advantage, that the reducing milieu within a cell leads to thecleavage of the linked PTD, and, thus, potential negative side effectsof the PTD on the function of the peptide of the present invention canbe avoided.

A further embodiment of the present invention relates to a nucleic acid,preferably a DNA, coding for a peptide of the present invention. Thenucleic acids coding for the peptides of the present invention can beplaced in expression vectors capable of expressing an encoded protein,polypeptide or peptide. Nucleic acids are inserted into vectors fromwhich they may be expressed by methods known to the person skilled inthe art. Vectors may, if desired, contain nucleic acids encodingportions of other proteins, thereby providing a fusion protein.

Therefore, the present invention is also directed to a recombinant DNAand an expression vector which includes any one of the present nucleicacids operably linked to regulatory control nucleic acid which effectsexpression of the nucleic acid in a host cell. The present invention isfurther directed to a host cell which contains such a recombinant DNA orsuch an expression vector.

Expression vectors include plasmids designed for the expression ofproteins or polypeptides fused to or within bacterial phage coatproteins. The DNA encoding the desired peptide, whether in a fusion,premature or mature form, may be ligated into expression vectorssuitable for any host. The DNA encoding the desired polypeptide may alsocontain a signal sequence to permit secretion from the intended host.Both prokaryotic and eukaryotic host systems are contemplated.

The present invention also contemplates a process for producing arecombinant peptide or immunogenic fragments thereof, encoded by anucleic acid of the present invention. The process involves: a)culturing a host cell which contains an expression vector having one ofthe nucleic acids of the present invention in a culture medium underconditions suitable for expression of one of said recombinant proteinsin the host cell, and b) isolating the recombinant protein from the hostcell or the culture medium.

The peptides of the present invention, whether in a premature, mature orfused form, are isolated from lysed cells, or from the culture medium,and are purified to the extent needed for the intended use. One of skillin the art can readily purify these proteins, polypeptides and peptidesby any available procedure. For example, purification may beaccomplished by salt fractionation, size exclusion chromatography, ionexchange chromatography, reverse phase chromatography, affinitychromatography and the like.

The present invention further contemplates antibodies which can bind tothe present peptides or fusion peptides, derivatives and fragmentsthereof. Such antibodies preferably bind to unique antigenic regions orepitopes in the peptides of the present invention. The antibody can bemonoclonal or polyclonal. To generate an antibody, animals, preferablyrabbits, chicken or mice are immunized with at least one peptide of thepresent invention, or a fragment or derivative thereof. The raisedantibodies can be isolated by well-known methods.

Epitopes and antigenic regions useful for generating antibodies can befound within the present peptides by procedures available to one ofskill in the art. For example, short, unique peptide sequences can beidentified in the present proteins and polypeptides that have little orno homology to known amino acid sequences. Preferably the region of aprotein selected to act as a peptide epitope or antigen is not entirelyhydrophobic; hydrophilic regions are preferred because those regionslikely constitute surface epitopes rather than internal regions of thepresent proteins and polypeptides. These surface epitopes are morereadily detected in samples tested for the presence of the presentproteins and polypeptides.

Peptides for immunization can be made by any procedure known to one ofskill in the art, for example, by using in vitro translation or chemicalsynthesis procedures. Short peptides which provide an antigenic epitopebut which by themselves are too small to induce an immune response maybe conjugated to a suitable carrier. Suitable carriers and methods oflinkage are well known in the art. Suitable carriers are typically largemacromolecules such as proteins, polysaccharides and polymeric aminoacids. Examples include serum albumins, keyhole limpet hemocyanin,ovalbumin, polylysine and the like. One of skill in the art can useavailable procedures and coupling reagents to link the desired peptideepitope to such a carrier. For example, coupling reagents can be used toform disulfide linkages or thioether linkages from the carrier to thepeptide of interest. If the peptide lacks a disulfide group, one may beprovided by the addition of a cysteine residue. Alternatively, couplingmay be accomplished by activation of carboxyl groups.

The minimum size of peptides useful for obtaining antigen specificantibodies can vary widely. The minimum size must be sufficient toprovide an antigenic epitope which is specific to the peptide. Themaximum size is not critical unless it is desired to obtain antibodiesto one particular epitope. For example, a large polypeptide may comprisemultiple epitopes, one epitope being particularly useful and a secondepitope being immunodominant.

The peptides of the present invention and/or their coding nucleic acidsare particularly useful to sensitize cells, particularly tumor cells,which are resistant to various treatment strategies to apoptosis.Whether the cells have been rendered to undergo apoptosis uponapplication of a peptide or a nucleic acid of the present invention canbe determined by a variety of methods known to the person skilled in theart, like for example TUNEL assay (TerminalDeoxynuceotidyltransferase-mediated UTP End Labelling), determination ofPARP cleavage by immunological methods, so-called caspase assays etc.

Furthermore, the peptides of the present invention and/or their codingnucleic acids, can be used as a pharmaceutical, optionally incombination with at least one active compound. This is a furtherembodiment of the present invention. The term “active compound” refersto a compound other than the peptide, a fragment or derivative thereof,which is able to induce apoptosis or which inhibits cell proliferation.

Active compounds which are able to induce apoptosis are known to theperson skilled in the art. One class of active compounds are chemicalcompounds having a cytostatic or antineoplastic effect (“cytostaticcompound”). Cytostatic compounds included in the present inventioncomprise, but are not restricted to (i) antimetabolites; (ii)DNA-fragmenting agents; (iii) DNA-crosslinking agents; (iv)intercalating agents; (v) protein synthesis inhibitors; (vi)topoisomerase I poisons; (vii) topoisomerase II poisons; (viii)microtubule-directed agents; (ix) kinase inhibitors; (x) miscellaneousinvestigational agents; (xi) hormones and (xii) hormone antagonists.

More specifically, the class of active compounds having a cytostatic oranti-neoplastic effect (“cytostatic compound”) as indicated aboveincluded in the present invention comprise, but are not restricted to(i) antimetabolites, such as cytarabine, fludarabine,5-fluoro-2′-deoxyuiridine, gemcitabine, hydroxyurea or methotrexate;(ii) DNA-fragmenting agents, such as bleomycin, (iii) DNA-crosslinkingagents, such as chlorambucil, cisplatin, cyclophosphamide or nitrogenmustard; (iv) intercalating agents such as adriamycin (doxorubicin) ormitoxantrone; (v) protein synthesis inhibitors, such as L-asparaginase,cycloheximide, puromycin or diphteria toxin; (vi) topoisomerase Ipoisons, such as camptothecin or topotecan; (vii) topoisomerase IIpoisons, such as etoposide (VP-16) or teniposide; (viii)microtubule-directed agents, such as colcemid, colchicine, paclitaxel,vinblastine or vincristine; (ix) kinase inhibitors such as flavopiridol,staurosporin, STI571 (CPG 57148B) or UCN-01 (7-hydroxystaurosporine);(x) miscellaneous investigational agents such as thioplatin, PS-341,phenylbutyrate, ET-18-OCH₃, or farnesyl transferase inhibitors(L-739749, L-744832); polyphenols such as quercetin, resveratrol,piceatannol, epigallocatechine gallate, theaflavins, flavanols,procyanidins, betulinic acid and derivatives thereof; (xi) hormones suchas glucocorticoids or fenretinide; (xii) hormone antagonists, such astamoxifen, finasteride or LHRH antagonists.

In a preferred embodiment of the present invention the cytostaticcompound is selected from the group consisting of cisplatin, doxorubicinand mitomycin C. Most preferred, the cytostatic compound is doxorubicin.

Another class of active compounds which can be used in the presentinvention are those which are able to sensitize for or induce apoptosisby binding to death receptors (“death receptor agonists”). Such agonistsof death receptors include death receptor ligands such as tumor necrosisfactor α (TNF-α), tumor necrosis factor β (TNF-β, lymphotoxin-α), LT-β(lymphotoxin-β), TRAIL (Apo2L, DR4 ligand), CD95 (Fas, APO-1) ligand,TRAMP (DR3, Apo-3) ligand, DR6 ligand as well as fragments andderivatives of any of said ligands. Preferably, the death receptorligand is TNF-α.

Furthermore, death receptors agonists comprise agonistic antibodies todeath receptors such as anti-CD95 antibody, anti-TRAIL-R1 (DR4)antibody, anti-TRAIL-R2 (DR5) antibody, anti-TRAIL-R3 antibody,anti-TRAIL-R4 antibody, anti-DR6 antibody, anti TNF-R1 antibody andanti-TRAMP (DR3) antibody as well as fragments and derivatives of any ofsaid antibodies.

For the purpose of sensitizing cells for apoptosis, the peptides and thenucleic acids of the present invention can be also used in combinationwith radiation therapy.

The phrase “radiation therapy” refers to the use of electromagnetic orparticulate radiation in the treatment of neoplasia. Radiation therapyis based on the principle that high-dose radiation delivered to a targetarea will result in the death of reproductive cells in both tumor andnormal tissues. The radiation dosage regimen is generally defined interms of radiation absorbed dose (rad), time and fractionation, and mustbe carefully defined by the oncologist. The amount of radiation apatient receives will depend on various consideration but the two mostimportant considerations are the location of the tumor in relation toother critical structures or organs of the body, and the extent to whichthe tumor has spread. Examples of radiotherapeutic agents are providedin, but not limited to, radiation therapy and is known in the art(Hellman, Principles of Radiation Therapy, Cancer, in Principles andPractice of Oncology, 24875 (Devita et al., 4^(th) ed., v1, 1993).Recent advances in radiation therapy include three-dimensional conformalexternal beam radiation, intensity modulated radiation therapy (IMRT),stereotactic radiosurgery and brachytherapy (interstitial radiationtherapy), the latter placing the source of radiation directly into thetumor as implanted “seeds”. These newer treatment modalities delivergreater doses of radiation to the tumor, which accounts for theirincreased effectiveness when compared to standard external beamradiation therapy.

Ionizing radiation with beta-emitting radionuclides is considered themost useful for radiotherapeutic applications because of the moderatelinear energy transfer (LET) of the ionizing particle (electron) and itsintermediate range (typically several millimeters in tissue). Gamma raysdeliver dosage at lower levels over much greater distances. Alphaparticles represent the other extreme; they deliver very high LETdosage, but have an extremely limited range and must, therefore, be inintimate contact with the cells of the tissue to be treated. Inaddition, alpha emitters are generally heavy metals, which limits thepossible chemistry and presents undue hazards from leakage ofradionuclide from the area to be treated. Depending on the tumor to betreated all kinds of emitters are conceivable within the scope of thepresent invention.

Furthermore, the present invention encompasses types of non-ionizingradiation like e.g. ultraviolet (UV) radiation, high energy visiblelight, microwave radiation (hyperthermia therapy), infrared (IR)radiation and lasers. In a particular embodiment of the presentinvention UV radiation is applied.

Generally, radiation therapy can be combined temporally with otheractive compounds listed above to improve the outcome of treatment. Thereare various terms to describe the temporal relationship of administeringradiation therapy together with other active compounds, and thefollowing examples are the preferred treatment regimens and aregenerally known by those skilled in the art and are provided forillustration only and are not intended to limit the use of othercombinations. Administration of radiation therapy with other activecompounds can be “sequential”, i.e. separately in time in order to allowthe separate administration, “concomitant” which refers to theadministration on the same day, and, finally, “alternating” which refersto the administration of radiation therapy on the days in which otheractive compounds would not have been administered.

A further object of the present invention are pharmaceuticalpreparations which comprise an effective dose of at least one of thedisclosed peptides and/or one of their coding nucleic acids, and/or atleast one active compound and a pharmaceutically acceptable carrier,i.e. one or more pharmaceutically acceptable carrier substances and/oradditives.

The dosage of the polypeptide or the nucleic acid, in combination withone or more active compounds to be administered, depends on theindividual case and is, as is customary, to be adapted to the individualcircumstances to achieve an optimum effect.

The peptides and their coding nucleic acids according to the presentinvention, respectively the medicaments containing the latter, can beused for the treatment of all cancer types which fail to undergoapoptosis. Examples of such cancer types comprise neuroblastoma,intestine carcinoma such as rectum carcinoma, colon carcinoma, familiaryadenomatous polyposis carcinoma and hereditary non-polyposis colorectalcancer, esophageal carcinoma, labial carcinoma, larynx carcinoma,hypopharynx carcinoma, tong carcinoma, salivary gland carcinoma, gastriccarcinoma, adenocarcinoma, medullary thyroidea carcinoma, papillarythyroidea carcinoma, renal carcinoma, kidney parenchym carcinoma,ovarian carcinoma, cervix carcinoma, uterine corpus carcinoma,endometrium carcinoma, chorion carcinoma, pancreatic carcinoma, prostatecarcinoma, testis carcinoma, breast carcinoma, urinary carcinoma,melanoma, brain tumors such as glioblastoma, astrocytoma, meningioma,medulloblastoma and peripheral neuroectodermal tumors, Hodgkin lymphoma,non-Hodgkin lymphoma, Burkitt lymphoma, acute lymphatic leukemia (ALL),chronic lymphatic leukemia (CLL), acute myeolid leukemia (AML), chronicmyeloid leukemia (CML), adult T-cell leukemia lymphoma, hepatocellularcarcinoma, gall bladder carcinoma, bronchial carcinoma, small cell lungcarcinoma, non-small cell lung carcinoma, multiple myeloma, basalioma,teratoma, retinoblastoma, choroidea melanoma, seminoma,rhabdomyosarcoma, craniopharyngeoma, osteosarcoma, chondrosarcoma,myosarcoma, liposarcoma, fibrosarcoma, Ewing sarcoma and plasmocytoma.

In a preferred embodiment of the invention, the cancer to be analyzed,diagnosed and treated is melanoma.

A further embodiment of the present invention is a diagnostic kitcomprising at least one peptide of the present invention, and/or anucleic acid encoding a peptide, a vector and a host cells.

The diagnostic kit can be used to detect IAPs, particularly livin, intumor cells that fail to undergo apoptosis, in particular melanoma.Methods to determine the presence and amount of IAPs in a given sampleare well known to the person skilled in the art. Briefly, a sample isprovided, said sample is contacted with a peptide that specificallybinds to livin and the presence or amount of the peptide bound to livinis determined, whereby the presence or amount of livin in said sample isdetermined. Methods to determine the amount and presence of peptidescomprise, among others, Western blotting, immunoprecipitation, ELISA,and RIA. For these purposes the peptides and nucleic acids of thepresent invention can be labelled with a suitable marker well-known tothe person skilled in the art.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1: Modified “Peptide Aptamer System” in S. cerevisiae.

Peptides of a peptide expression library with a length of 20 amino acidsare co-expressed as a fusion peptides fused to a transcriptionaltransactivator domain (GAL4AD), together with the target protein X (e.g.livin) which is fused to a DNA binding domain (GAL4BD). Upon successfulbinding of a peptide with the target protein, a synthetic transcriptionfactor is created which in turn binds to the binding site and activatesthe transcription of a selection marker (e.g. ADE2). Using selectivegrowth conditions, e.g. lacking adenine on growth media, only thoseyeast colonies are able to grow, which express a peptide that binds tothe target (NLS: nuclear localization signal; TAG: suitable marker todetect the peptide aptamer; trxA: E. coli Thioredoxin A scaffold)

FIG. 2: Interaction in Mammalian Cells

The FIG. 2 shows the interaction results from a mammalianTwo-Hybrid-System (CheckMate, Promega, Madison Wis.). The Y-axisdisplays relative luciferase activity of the peptides of the presentinvention (particularly SEQ ID NOs: 87-132). The differently stainedbars represent independent CheckMate analyses.

FIG. 3: Colony Formation Assay

Expression vectors bearing the nucleic acid coding for peptide number 75(corresponds to SEQ ID No: 127 or number 41 of TABLE 2) or bearing thenucleic acid coding for control peptide were introduced in HeLa cellsand livin-negative C33a cells. The cells were selected for stabletransfectants.

FIG. 4: Induction of Apoptosis

Peptide number 75 as defined for FIG. 3, linked to poly-arginine R9, wasadded to the cell culture medium of HeLa cells and livin-negative H1299cells. Upper panel left of FIG. 4 shows the scheme of how cells weretreated with the cytostatic compound doxorubicin (0.5 μg/l ml). Upperpanel right shows Western Blot of active caspase 3 and cleaved PARP.Lower panel shows increase of apoptotic cells.

TABLE 1: Sequences of Peptides

The Table displays the sequences of the 86 peptides of the presentinvention (NS=nucleotide sequence; AS=amino acid sequence; *=break ofpeptide chain, resulting in a “linear peptide” fused to the N-terminalpart of trxA; ND=Not Determined))

TABLE 2: Sequences of Linear Peptides

Table 2 lists so-called linear peptides being not embedded in athioredoxin A scaffold as it is commonly done for aptamers. Linearpeptide banks are generated by expressing randomized 20-mer peptidesduring a screening procedure as GAL4AD fusions (i.e. aminoterminallyfused to the GAL4 activation domain). This results in peptides with afree carboxyterminal end.

TABLE 3: Specificity for Livin

Table 3 lists the peptides with high specificity to livin versus otherIAPs. The numbers of the peptides in the first column reflects theinventors' numbering (5=SEQ ID:90, 7=SEQ ID:91, 15=SEQ ID:98, 16=SEQID:99, 40=SEQ ID:111, 54=SEQ ID: 18, 65=SEQ ID123, 69=SEQ ID:124, 75=SEQID 127, 77=SEQ ID:128, 90=SEQ ID:132). K1 refers to a control peptide.Livin-β, c-IAP-1, c-LAP-2, XIAP and surivin refer to different IAPsbeing tested for binding. HPV16 E6 refers to human papilloma virus type16 E6 protein being used as a control.

EXAMPLES Example 1 Screening for Livin-Binding Peptides

A method derived from the “peptide aptamer system” is used. Yeast strainKF1 (MATa trp1-901 leu2-3,112 his 3-200 gal4Δ gal80Δ LYS2::GAL1-HIS3GAL2-ADE2 met2::GAL7-lacZ SPAL10-URA3) is generated from PJ69-4A afterintegration at the ura3-52 locus by homologous recombination of PCRproduct encompassing the SPAL10-URA3 allele from yeast strain MaV103.KF1 thus contains three selectable marker genes under the control ofGal4-binding sites: GAL2-ADE2, GAL1-HIS3 and SPO13-URA3. As bait, thecomplete livin sequence is fused in frame to the Gal4 DNA-binding domaininto vector pPC97, yielding pPC97/livin. A yeast expression vector,pADtrx, in which the ADH1 promoter directs the expression the E. colithioredoxin A (trxA) fused to the Gal4 activation domain, is constructedfrom pRS424. In addition, pADtrx contains the simian virus 40 nuclearlocalization signal and an influenza hemagglutinin epitope. A randomizedpeptide expression library is generated in pADtrx by cloning randomized60-mer oligonucleotides into the unique RsrII site of trxA.Oligonucleotides contained triplets of the sequence NKK (where N=G, A, Tor C and K=G or C), which encode for all 20 amino acids but result inonly one stop codon. The complexity of the peptide aptamer expressionlibrary is estimated to be in the range of 2×10⁸ different members.

KF1 transformants expressing pPC97/livin and the peptide aptamerexpression library are selected initially for growth in the absence ofadenine. Subsequently, they are analyzed by replica plating foractivation of the Gal4-dependent GAL2-ADE2, GAL1-HIS3 and SPO13-URA3genes. Peptide aptamer expression vectors from clones exhibiting growthin the absence of adenine and histidine are rescued, and activation ofthe selectable markers is verified by rescreening. Binding specificityof livin binding aptamers was tested by using control proteins as baitslike HPV16 E6 or HPV16E7.

Using the above method, 38 peptides have been identified that in thecontext of trxA specifically interact with livin (see Table 1 SEQ IDNOS: 1-38). Alternatively, a peptide aptamer library was directly fusedto the Gal4 activation domain, giving rise to “linear” peptides that arenot embedded in the trxA scaffold. Using this approach, another 48peptides that specifically interact with livin have been identified(Table 1 SEQ ID NOS:39-86).

Peptides according to SEQ ID NOs:87-132 are also so-called “linear”peptides, not being embedded in a TrxA scaffold.

Example 2 Mammalian Two-Hybrid Analyses

In order to analyze the aptamer/livin interactions in human cells, the“CheckMate™ Mammalian Two-Hybrid System” (Promega GmbH, Mannheim,Germany) is employed. The complete livin protein was expressed as a Gal4binding domain fusion protein from pBIND. Individual peptide aptamersare expressed within the trxA scaffold, linked to the transcriptionalactivation domain of the herpes simplex virus-1 VP16 protein, from pACT.Reporter plasmid pG5luc contains the firefly luciferase under thetranscriptional control of five Gal4-binding sites. Cells are harvested24 hours after transfection, and luciferase activities are measured bymethods known to the person skilled in the art.

Using this approach, it is shown that the interaction of the selectedpeptides to livin occurs also in mammalian cells.

As shown in FIG. 2, the majority of linear petides according to TABLE 2(SEQ ID NOs: 87-132), interact in mammalian cells with livin, asverified by a high luciferase activity.

Example 3 Specificity for Livin

For further analyses in the yeast two hybrid system, i.e. theinteraction with other IAPs that livin, 11 peptides (see TABLE 3) whichexerted the highest activity in the mammalian Two-Hybrid-System werechosen. As depicted in Table 3, the peptides of the present inventionbind in a highly specific manner to Livin (beta) and not or only to aneglectible extent to other IAPs. This is particularly advantageous forthe use of the peptides for therapeutic purposes, since livin is highlyexpressed only in tumor cells. Other IAPs may fulfil physiologicfunctions in non-tumorous cells. Thus, a binding and inactivation ofthese IAPs can result in undesired side effects.

Example 4 Livin-Binding Peptides Block Growth of Livin-Binding HeLaCells

The nucleic acids coding for the peptides of TABLE 3 were cloned intomammalian expression vectors and transfected into different cells toobserve their further growth. By using a HIS-tag it was possible toverify that all peptide sequences could be expressed. As depicted inFIG. 3, most of the peptides, in particular peptide number 75, blockedthe growth of livin-expressing HeLa and MeWo (melanoma cells), whereaslivin-negative cells like C33a were not affected.

Example 5 Livin-Binding Peptide Fused to a Carrier SensitizesLivin-Positive Cells for Apoptosis

A synthetic peptide by fusing peptide number 75 (SEQ ID NO:127) topoly-arginine R9 was generated by methods known to the person skilled inthe art. After adding the fusion peptide to the culture medium, anefficient intracellular uptake of the peptide could be verified by usinga HIS-tag (100% of cells within 20 min). The ectopic expression of thepeptide leaded to a sensitization of livin-positive cells forpro-apoptotic drugs, e.g. cytostatic compounds like doxorubicin). Therepeated addition of doxorubicin resulted in a increased concentrationof active caspase as shown in FIG. 4. The control peptide sc75(scrambled form of peptide number 75 with randomised amino acidsequence) did not have this effect. The increased caspase-3 activitydirectly correlated with an increased cleavage of the caspase-3substrate PARP and an increase of apoptosis of HeLa but not of H1299cells. TABLE 1 SEQUENCE IDENTIFICATION NUMBERS SEQ ID NO NAME NS ASSequence 1 LT 1-1 60 20 WLGTFSGTCSTAFYFPLGVP 2 LT 6-1 60 20CRWLRTKRTLPLFSVMPFWC 3 LT 7-1 61 31 MYSNVSVDVAADGVSCVCCSWSVQNDRPDSG* 4LT 8-2 60 20 YKWRMGVYLSGVRLMRAFII 5 LT 9-1 60 20 VSYRTCTAGGQMSRWRLFII 6LT 12-1 60 20 GYSLTSMSAFAVRPCVCGSL 7 LT 13-1 60 20 WLGTFSGTCSTAFYFPLGVP8 LT 15-2 60 20 TNFRPSPTFHAILLWPNTFS 9 LT 16-1 60 20VGLGGWCFDCYWVAWDFQTQ 10 LT 19-2 60 20 FWDYCGPLICLHGNLGRCVS 11 LT 21-2 61 5 MMDSA* 12 LT 22-2 60 20 ASLRLYPIGGTVPFGRTGAG 13 LT 23-2 60 20GDYGCCWVVTTGVGVRCYVW 14 LT 27-1 60 20 ACWALWSLFRQDLLLVITFD 15 LT 28-1 6131 DSAPGERYFVDFLGVSFACVWSVQNDRPDSG* 16 LT 29-1 60 14 IPWAPPMYFADSNV* 17LT 33-1 60 20 TPSCRAGVLRCTGCFGVRSG 18 LT 34-2 60 20 LWRGRTVSAYLSWLRHYSSS19 LT 35-1 60 20 HSRPALCMVSLRWARSLWIV 20 LT 37-2 42 14+? WTHVWVGWLVAGMS21 LT 38-1 60 20 RFRCRADLCVTLTVLSFLAQ 22 LT 45-2 60 20CLETLRVCPYVARIAIQHLR 23 LT 48-3 60 20 LLAWRVQQSRPLPYLHIAFI 24 LT 49-2 6020 PPPLTGRWSRQCVSVFGIVH 25 LT 54-1 60 20 CWIHRAWMLSWHGVWSLTLV 26 LT 62-160 20 APPISGRWRGLYMRSRFVSL 27 LT 76-3 60 18 VRLFIVCIIICCLMLLVG* 28 LT79-1 60 20 IPSCSVLVCLCHLARLWHCE 29 LT 82-3 60 20 CSVMHVFRVGPGSSGSLSCG 30LT 93-1 126 42 RATYWFRSRYQVVHRSRLPYGPLIVVGIGALNLELNRTLLCS 31 LT 103-1 6020 SLAIWSTQSCARCQCTLSRV 32 LT 105-2 60 20 FWFLPAPPCKCGLLYRLSVH 33 LT108-1 60 20 LAGRHFSRVVDRIRYRLLWT 34 LT 126-2 60 20 MPIPPLCRSAGRLLYLYTHY35 LT 127-1 60 20 YTLPSVLLCLMRTGMLRCAC 36 LT 135-2 60 20GMPIRASPCYLGVDGWCXTL 37 LT 138-2 60 20 KPWEYLRMFPWMRVARFFIW 38 LT 140-260 20 ALLMFGCPNWFASWRLHLFI 39 ND ND NDEFGSGLVSGRGIIVRRMLFLRVLLWVLRSAEIYES* 40 ND ND NDGPFENWRVEELARGRYRMHGDVVLRSAEIYES* 41 ND ND ND GPIDCIIFLLWYSRQQRGGSRGGP*42 ND ND ND GSRFRVFVCSLFSFLSGRGGGVVVLRSAEIYES* 43 ND ND NDGPFKRCHERLVAFARCWFMWSMVLRSAEIYES* 44 ND ND NDGPSNDNQLVLRVRILRVLIVMRVLRSAEIYES* 45 ND ND NDEFRVRRMRLLVRLMGSDDSGTIPDFGP* 46 ND ND ND GPSLQFLEVVVSCYMVLYDLSKGP* 47 NDND ND GPQPFCSPPSFYTRLLIIVRLLSLDLQRSSNRRY* 48 ND ND NDGPAPLSLCVCKCGCGHTRPFVGP* 49 ND ND ND GPDVHIWQSIIFYAMRHMMGP* 50 ND ND NDEFGSGCGCFVRGRIVRIRCVILLLRVLRSAEIYES* 51 ND ND NDGPHSSAHDRIWLRVRGLRIILLVLRSAEIYES* 52 ND ND NDEFGSGLCVRRWWGMSVGSRIMLVMLVLRSAEIYES* 53 ND ND NDGPVYSEAFVCLVCAGVCVEECGGSLDLQRSSNRRY* 54 ND ND NDGPIETVGFIVRLHTLLMVLRRTGP* 55 ND ND ND GPLHRTLLVDMCCWLMSLESNMGP* 56 ND NDND EFGVRVVCVVRSLFVLRCGLLRCRGVLRSAEIYES* 57 ND ND NDEFVRECSLCRVMVLMFVLRGIRLRVLRSAEIYES* 58 ND ND NDEFGVRLLVLLRLRCVRRGGGCFVCWVLRSAEIYES* 59 ND ND NDGSGFRMRVLVMVCRLRVVFLVRRVLRSAEIYES* 60 ND ND NDGRLGWLRLLCVRIVLVCLRRGLVLRSAEIYES* 61 ND ND NDEFGSGWYVDLGDYSVWVDYVYCGSGSLDLQRSSNRRY* 62 ND ND NDGPSQCAQRVALIQMYIDALVCIGP* 63 ND ND NDEFGSGCVRIRVGIVRRMLXLRFVFLVLRSAEIYES* 64 ND ND NDGPLLDPMLRQRFSLRVWIMLLGVLRSAEIYES* 65 ND ND ND GPNSWVWRYVTIAHWLANYRMSGP*66 ND ND ND GPAMKSCTIRVFRVCIVLRIVRVLRSAEIYES* 67 ND ND NDEFGVRMRLMIRIFRGLFVLRGFRGLVLRSAEIYES* 68 ND ND NDGPVPSSPCSFLLYCRDVLCHWPGP* 69 ND ND ND GPCEPFIGDCWPCLIRTLVTLRGLDL 70 NDND ND GPWWKDRGVLVRLCVLRLVVGVVLRSAEIYES* 71 ND ND NDGPRLLVRMRGWCRVSLIXFWLRVLRSAEIYES* 72 ND ND NDEFRVRIIVVSLRIWRLLVRRRCLCLVLRSAEIYES* 73 ND ND NDGPVECADVLFASRIRLLCLCFRVLRSAEIYES* 74 ND ND NDEFGRRLLVFRLSVFVVVLGRRLSRVLRSAEIYES* 75 ND ND NDGAGLGRVIRLRIVVLRCIFLLFRVLRSAEIYES* 76 ND ND NDGPFPFDYPRWIMIVLLRGVLRSAEIYES* 77 ND ND NDGSRGLRLCLLGRCRLCGCLIIMRVLRSAEIYES* 78 ND ND ND GPESYVLWPARGEALYYLRAWLGP*79 ND ND ND GSRCIRRRISILFFVFRVLRSRRVLRSAEIYES* 80 ND ND NDGPFSEHARGHVVTICRLRLLFWLLRSAEIYES* 81 ND ND NDGPSSLLRRCLILGMVLGVLRRRVLRSAEIYES* 82 ND ND ND GPHPVLAVQLINAYLGLERVGRGP*83 ND ND ND GPLPSGAVSTEAYFWEVFKLLMGP* 84 ND ND NDGPYPYLRILLVQKIACVRRALWVLRSAEIYES* 85 ND ND NDGPVGVEGVDSVFGWCVVCFLLVWSLDLQRSSNRRY* 86 ND ND NDEFRVRVLGCMGVFLRLRFCGGLRLRVLRSAEIYES*

TABLE 2 Number Peptide Sequence 1 1 GSGLVSGRGIIVRRMLFLRVLLWVLRSAEIYES* 23 GPFENWRVEELARGRYRMHGDVVLRSAEIYES* 3 4 GPIDCIIFLLWYSRQQRGGSRGGP* 4 5GSRFRVFVCSLFSFLSGRGGGVVVLRSAEIYES* 5 7 GPFKRCHERLVAFARCWFMWSMVLRSAEIYES*6 8 GPSNDNQLVLRVRILRVLIVMRVLRSAEIYES* 7 9 RVRRMRLLVRLMGSDDSGTIPDFGP* 810 GPSLQFLEVVVSCYMVLYDLSKGP* 9 11 GPQPFCSPPSFYTRLLIIVRLLSLDLQRSSNRRY* 1013 GPAPLSLCVCKCGCGHTRPFVGP* 11 14 GPDVHIWQSIIFYAMRHMMGP* 12 15GSGCGCFVRGRIVRIRCVILLLRVLRSAEIYES* 13 16GPHSSAHDRIWLRVRGLRIILLVLRSAEIYES* 14 17GSGLCVRRWWGMSVGSRIMLVMLVLRSAEIYES* 15 18GPVYSEAFVCLVCAGVCVEECGGSLDLQRSSNRRY* 16 19 GPIETVGFIVRLHTLLMVLRRTGP* 1720 GPLHRTLLVDMCCWLMSLESNMGP* 18 22 GVRVVCVVRSLFVLRCGLLRCRGVLRSAEIYES* 1923 VRECSLCRVMVLMFVLRGIRLRVLRSAEIYES* 20 28GVRLLVLLRLRCVRRGGGCFVCWVLRSAEIYES* 21 30GSGFRMRVLVMVCRLRVVFLVRRVLRSAEIYES* 22 33GRLGWLRLLCVRIVLVCLRRGLVLRSAEIYES* 23 34GSGWYVDLGDYSVWVDYVYCGSGSLDLQRSSNRRY* 24 37 GPSQCAQRVALIQMYIDALVCIGP* 2540 GSGCVRIRVGIVRRMLFLRFVFLVP* 26 41 GPLLDPMLRQRFSLRVWIMLLGVLRSAEIYES* 2748 GPNSWVWRYVTIAHWLANYRMSGP* 28 49 GPAMKSCTIRVFRVCIVLRIVRVLRSAEIYES* 2951 GVRMRLMIRIFRGLFVLRGFRGLVLRSAEIYES* 30 52 GPVPSSPCSFLLYCRDVLCHWPGP* 3153 GPCEPFIGDCWPCLIRTLVTLRGLDLQRSSNRRY* 32 54GPWWKDRGVLVRLCVLRLVVGVVLRSAEIYES* 33 57GPRLLVRMRGWCRVSLIXFWLRVLRSAEIYES* 34 59RVRIIVSLRIWRLLVRRRCLCLVLRSAEIYES* 35 60GPVECADVLFASRIRLLCLCFRVLRSAEIYES* 36 62GRRLLVFRLSVFVVVLGRRLSRVLRSAEIYES* 37 65GAGLGRVIRLRIVVLRCIFLLFRVLRSAEIYES* 38 69 GPFPFDYPRWIMIVLLRGVLRSAEIYES*39 72 GSRGLRLCLLGRCRLCGCLIIMRVLRSAEIYES* 40 74 GPESYVLWPARGEALYYLRAWLGP*41 75 GSRCIRRRISILFFVFRVLRSRRVLRSAEIYES* 42 77GPFSEHARGHVVTICRLRLLFWLLRSAEIYES* 43 79GPSSLLRRCLILGMVLGVLRRRVLRSAEIYES* 44 88 GPHPVLAVQLINAYLGLERVGRGP* 45 89GPLPSGAVSTEAYFWEVFKLLMGP* 46 90 GPYPYLRILLVQKIACVRRALWVLRSAEIYES*

TABLE 3 (linear) peptide Livinβ c-IAP-1 c-IAP-2 XIAP Survivin HPV16 E6 5 + − − − − −  7 + − − − − − 15 + − − − − − 16 + − − − − − 40 + − − − −− 54 + − − − − − 65 + − − − − − 69 + − − − − − 75 + − − − − − 77 + − − +− − 90 + − − − − − K1 − − − − − −

1-21. (canceled)
 22. A peptide, a fragment or derivative thereof, whichsensitizes cells for apoptosis, comprising an amino acid sequenceselected from the group of SEQ ID NOS:1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29,30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47,48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65,66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83,84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100,101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114,114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127,128, 129, 130, 131,
 132. 23. A peptide according to claim 22, whereinthe peptide is linked to a second moiety which mediates the uptake intoa cell.
 24. A peptide according to claim 23, wherein the second moietyis a carrier.
 25. A nucleic acid coding for a peptide as defined inclaim
 22. 26. A recombinant DNA comprising at least one nucleic acidcoding for a peptide as defined in claim 22, operably linked toregulatory control nucleic acid sequences which can affect expression ofsaid nucleic acid sequences in a host cell.
 27. An expression vector orplasmid comprising the recombinant DNA as defined in claim
 26. 28. Ahost cell comprising the expression vector of claim
 27. 29. The hostcell according to claim 28, wherein the host cell is a eukaryotic orprokaryotic cell.
 30. A method of producing a peptide or an immunogenicfragment thereof, encoded by the recombinant DNA according to claim 26,which process comprises (i) culturing a host cell in a culture mediumsuitable for the expression of said proteins or immunogenic fragmentsthereof, and (ii) recovering said recombinant proteins or immunogenicfragments thereof from said host cell or said culture medium.
 31. Anantibody that immunospecifically binds to a peptide according as definedin claim
 22. 32. A method of sensitizing a cell for apoptosis by usingat least one peptide of claim 22, optionally in combination with atleast one active compound.
 33. A method of sensitizing a cell forapoptosis by using a nucleic acid of claim 25, optionally in combinationwith at least one active compound.
 34. The method according to claim 32,wherein the sensitization for apoptosis occurs by binding of IAPs. 35.The method according to claim 34, wherein the IAP is livin-β.
 36. Themethod according to claim 35, wherein the cell which is sensitized forapoptosis is a cancer cell.
 37. The method according to claim 36,wherein the cancer cell is selected from the group consisting ofneuroblastoma, intestine carcinoma preferably rectum carcinoma, coloncarcinoma, familiary adenomatous polyposis carcinoma and hereditarynon-polyposis colorectal cancer, esophageal carcinoma, labial carcinoma,larynx carcinoma, hypopharynx carcinoma, tong carcinoma, salivary glandcarcinoma, gastric carcinoma, adenocarcinoma, medullary thyroideacarcinoma, papillary thyroidea carcinoma, renal carcinoma, kidneyparenchym carcinoma, ovarian carcinoma, cervix carcinoma, uterine corpuscarcinoma, endometrium carcinoma, chorion carcinoma, pancreaticcarcinoma, prostate carcinoma, testis carcinoma, breast carcinoma,urinary carcinoma, melanoma, brain tumors preferably glioblastoma,astrocytoma, meningioma, medulloblastoma and peripheral neuroectodermaltumors, Hodgkin lymphoma, non-Hodgkin lymphoma, Burkitt lymphoma, acutelymphatic leukemia (ALL), chronic lymphatic leukemia (CLL), acutemyeolid leukemia (AML), chronic myeloid leukemia (CML), adult T-cellleukemia lymphoma, hepatocellular carcinoma, gall bladder carcinoma,bronchial carcinoma, small cell lung carcinoma, non-small cell lungcarcinoma, multiple myeloma, basalioma, teratoma, retinoblastoma,choroidea melanoma, seminoma, rhabdomyosarcoma, craniopharyngeoma,osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma,Ewing sarcoma and plasmocytoma.
 38. The method according to claim 37,wherein the cancer is melanoma.
 39. A medicament for the treatment ofcancer, comprising a peptide of claim 22, and a pharmaceuticallyacceptable carrier, optionally in combination with an active compound.40. The medicament according to claim 39, wherein the cancer is selectedfrom the group consisting of neuroblastoma, intestine carcinomapreferably rectum carcinoma, colon carcinoma, familiary adenomatouspolyposis carcinoma and hereditary non-polyposis colorectal cancer,esophageal carcinoma, labial carcinoma, larynx carcinoma, hypopharynxcarcinoma, tong carcinoma, salivary gland carcinoma, gastric carcinoma,adenocarcinoma, medullary thyroidea carcinoma, papillary thyroideacarcinoma, renal carcinoma, kidney parenchym carcinoma, ovariancarcinoma, cervix carcinoma, uterine corpus carcinoma, endometriumcarcinoma, chorion carcinoma, pancreatic carcinoma, prostate carcinoma,testis carcinoma, breast carcinoma, urinary carcinoma, melanoma, braintumors preferably glioblastoma, astrocytoma, meningioma, medulloblastomaand peripheral neuroectodermal tumors, Hodgkin lymphoma, non-Hodgkinlymphoma, Burkitt lymphoma, acute lymphatic leukemia (ALL), chroniclymphatic leukemia (CLL), acute myeolid leukemia (AML), chronic myeloidleukemia (CML), adult T-cell leukemia lymphoma, hepatocellularcarcinoma, gall bladder carcinoma, bronchial carcinoma, small cell lungcarcinoma, non-small cell lung carcinoma, multiple myeloma, basalioma,teratoma, retinoblastoma, choroidea melanoma, seminoma,rhabdomyosarcoma, craniopharyngeoma, osteosarcoma, chondrosarcoma,myosarcoma, liposarcoma, fibrosarcoma, Ewing sarcoma and plasmocytoma.41. The medicament according to claim 40, wherein the active compound isselected from the group consisting of (i) antimetabolites; (ii)DNA-fragmenting agents; (iii) DNA-crosslinking agents; (iv)intercalating agents; (v) protein synthesis inhibitors; (vi)topoisomerase I poisons; (vii) topoisomerase II poisons; (viii)microtubule-directed agents; (ix) kinase inhibitors; (x) miscellaneousinvestigational agents; (xi) hormones and (xii) hormone antagonists. 42.A medicament for the treatment of cancer, comprising a nucleic acid ofclaim 25, and a pharmaceutically acceptable carrier, optionally incombination with an active compound.
 43. The medicament according toclaim 42, wherein the cancer is selected from the group consisting ofneuroblastoma, intestine carcinoma preferably rectum carcinoma, coloncarcinoma, familiary adenomatous polyposis carcinoma and hereditarynon-polyposis colorectal cancer, esophageal carcinoma, labial carcinoma,larynx carcinoma, hypopharynx carcinoma, tong carcinoma, salivary glandcarcinoma, gastric carcinoma, adenocarcinoma, medullary thyroideacarcinoma, papillary thyroidea carcinoma, renal carcinoma, kidneyparenchym carcinoma, ovarian carcinoma, cervix carcinoma, uterine corpuscarcinoma, endometrium carcinoma, chorion carcinoma, pancreaticcarcinoma, prostate carcinoma, testis carcinoma, breast carcinoma,urinary carcinoma, melanoma, brain tumors preferably glioblastoma,astrocytoma, meningioma, medulloblastoma and peripheral neuroectodermaltumors, Hodgkin lymphoma, non-Hodgkin lymphoma, Burkitt lymphoma, acutelymphatic leukemia (ALL), chronic lymphatic leukemia (CLL), acutemyeolid leukemia (AML), chronic myeloid leukemia (CML), adult T-cellleukemia lymphoma, hepatocellular carcinoma, gall bladder carcinoma,bronchial carcinoma, small cell lung carcinoma, non-small cell lungcarcinoma, multiple myeloma, basalioma, teratoma, retinoblastoma,choroidea melanoma, seminoma, rhabdomyosarcoma, craniopharyngeoma,osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma,Ewing sarcoma and plasmocytoma.
 44. A medicament according to claim 43,wherein the active compound is selected from the group consisting of (i)antimetabolites; (ii) DNA-fragmenting agents; (iii) DNA-crosslinkingagents; (iv) intercalating agents; (v) protein synthesis inhibitors;(vi) topoisomerase I poisons; (vii) topoisomerase II poisons; (viii)microtubule-directed agents; (ix) kinase inhibitors; (x) miscellaneousinvestigational agents; (xi) hormones and (xii) hormone antagonists. 45.A medicament for the treatment of cancer, comprising a vector of claim27, and a pharmaceutically acceptable carrier, optionally in combinationwith an active compound.
 46. A medicament according to claim 45, whereinthe cancer is selected from the group consisting of neuroblastoma,intestine carcinoma preferably rectum carcinoma, colon carcinoma,familiary adenomatous polyposis carcinoma and hereditary non-polyposiscolorectal cancer, esophageal carcinoma, labial carcinoma, larynxcarcinoma, hypopharynx carcinoma, tong carcinoma, salivary glandcarcinoma, gastric carcinoma, adenocarcinoma, medullary thyroideacarcinoma, papillary thyroidea carcinoma, renal carcinoma, kidneyparenchym carcinoma, ovarian carcinoma, cervix carcinoma, uterine corpuscarcinoma, endometrium carcinoma, chorion carcinoma, pancreaticcarcinoma, prostate carcinoma, testis carcinoma, breast carcinoma,urinary carcinoma, melanoma, brain tumors preferably glioblastoma,astrocytoma, meningioma, medulloblastoma and peripheral neuroectodermaltumors, Hodgkin lymphoma, non-Hodgkin lymphoma, Burkitt lymphoma, acutelymphatic leukemia (ALL), chronic lymphatic leukemia (CLL), acutemyeolid leukemia (AML), chronic myeloid leukemia (CML), adult T-cellleukemia lymphoma, hepatocellular carcinoma, gall bladder carcinoma,bronchial carcinoma, small cell lung carcinoma, non-small cell lungcarcinoma, multiple myeloma, basalioma, teratoma, retinoblastoma,choroidea melanoma, seminoma, rhabdomyosarcoma, craniopharyngeoma,osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma,Ewing sarcoma and plasmocytoma.
 47. A medicament according to claim 46,wherein the active compound is selected from the group consisting of (i)antimetabolites; (ii) DNA-fragmenting agents; (iii) DNA-crosslinkingagents; (iv) intercalating agents; (v) protein synthesis inhibitors;(vi) topoisomerase I poisons; (vii) topoisomerase II poisons; (viii)microtubule-directed agents; (ix) kinase inhibitors; (x) miscellaneousinvestigational agents; (xi) hormones and (xii) hormone antagonists. 48.A method for the treatment of cancer, the method comprisingadministering to a patient a medicament of claim
 39. 49. The methodaccording to claim 48, wherein the cancer is selected from the groupconsisting of neuroblastoma, intestine carcinoma preferably rectumcarcinoma, colon carcinoma, familiary adenomatous polyposis carcinomaand hereditary non-polyposis colorectal cancer, esophageal carcinoma,labial carcinoma, larynx carcinoma, hypopharynx carcinoma, tongcarcinoma, salivary gland carcinoma, gastric carcinoma, adenocarcinoma,medullary thyroidea carcinoma, papillary thyroidea carcinoma, renalcarcinoma, kidney parenchym carcinoma, ovarian carcinoma, cervixcarcinoma, uterine corpus carcinoma, endometrium carcinoma, chorioncarcinoma, pancreatic carcinoma, prostate carcinoma, testis carcinoma,breast carcinoma, urinary carcinoma, melanoma, brain tumors preferablyglioblastoma, astrocytoma, meningioma, medulloblastoma and peripheralneuroectodermal tumors, Hodgkin lymphoma, non-Hodgkin lymphoma, Burkittlymphoma, acute lymphatic leukemia (ALL), chronic lymphatic leukemia(CLL), acute myeolid leukemia (AML), chronic myeloid leukemia (CML),adult T-cell leukemia lymphoma, hepatocellular carcinoma, gall bladdercarcinoma, bronchial carcinoma, small cell lung carcinoma, non-smallcell lung carcinoma, multiple myeloma, basalioma, teratoma,retinoblastoma, choroidea melanoma, seminoma, rhabdomyosarcoma,craniopharyngeoma, osteosarcoma, chondrosarcoma, myosarcoma,liposarcoma, fibrosarcoma, Ewing sarcoma and plasmocytoma.
 50. A methodfor the treatment of cancer, the method comprising administering topatient a medicament of claim
 42. 51. The method according to claim 50,wherein the cancer is selected from the group consisting ofneuroblastoma, intestine carcinoma preferably rectum carcinoma, coloncarcinoma, familiary adenomatous polyposis carcinoma and hereditarynon-polyposis colorectal cancer, esophageal carcinoma, labial carcinoma,larynx carcinoma, hypopharynx carcinoma, tong carcinoma, salivary glandcarcinoma, gastric carcinoma, adenocarcinoma, medullary thyroideacarcinoma, papillary thyroidea carcinoma, renal carcinoma, kidneyparenchym carcinoma, ovarian carcinoma, cervix carcinoma, uterine corpuscarcinoma, endometrium carcinoma, chorion carcinoma, pancreaticcarcinoma, prostate carcinoma, testis carcinoma, breast carcinoma,urinary carcinoma, melanoma, brain tumors preferably glioblastoma,astrocytoma, meningioma, medulloblastoma and peripheral neuroectodermaltumors, Hodgkin lymphoma, non-Hodgkin lymphoma, Burkitt lymphoma, acutelymphatic leukemia (ALL), chronic lymphatic leukemia (CLL), acutemycolid leukemia (AML), chronic myeloid leukemia (CML), adult T-cellleukemia lymphoma, hepatocellular carcinoma, gall bladder carcinoma,bronchial carcinoma, small cell lung carcinoma, non-small cell lungcarcinoma, multiple myeloma, basalioma, teratoma, retinoblastoma,choroidea melanoma, seminoma, rhabdomyosarcoma, craniopharyngeoma,osteosarcoma, chondrosarcoma, myosarcoma, liposarcoma, fibrosarcoma,Ewing sarcoma and plasmocytoma.
 52. A method for the treatment ofcancer, the method comprising to a patient a medicament of claim
 45. 53.The method according to claim 52, wherein the cancer is selected fromthe group consisting of neuroblastoma, intestine carcinoma preferablyrectum carcinoma, colon carcinoma, familiary adenomatous polyposiscarcinoma and hereditary non-polyposis colorectal cancer, esophagealcarcinoma, labial carcinoma, larynx carcinoma, hypopharynx carcinoma,tong carcinoma, salivary gland carcinoma, gastric carcinoma,adenocarcinoma, medullary thyroidea carcinoma, papillary thyroideacarcinoma, renal carcinoma, kidney parenchym carcinoma, ovariancarcinoma, cervix carcinoma, uterine corpus carcinoma, endometriumcarcinoma, chorion carcinoma, pancreatic carcinoma, prostate carcinoma,testis carcinoma, breast carcinoma, urinary carcinoma, melanoma, braintumors preferably glioblastoma, astrocytoma, meningioma, medulloblastomaand peripheral neuroectodermal tumors, Hodgkin lymphoma, non-Hodgkinlymphoma, Burkitt lymphoma, acute lymphatic leukemia (ALL), chroniclymphatic leukemia (CLL), acute myeolid leukemia (AML), chronic myeloidleukemia (CML), adult T-cell leukemia lymphoma, hepatocellularcarcinoma, gall bladder carcinoma, bronchial carcinoma, small cell lungcarcinoma, non-small cell lung carcinoma, multiple myeloma, basalioma,teratoma, retinoblastoma, choroidea melanoma, seminoma,rhabdomyosarcoma, craniopharyngeoma, osteosarcoma, chondrosarcoma,myosarcoma, liposarcoma, fibrosarcoma, Ewing sarcoma and plasmocytoma.54. A diagnostic kit for the detection of IAPs, preferably livin-β, incancer cells, comprising at least one peptide as defined in claim 22.55. A diagnostic kit for the detection of IAPs, preferably livin-β,preferably in cancer cells, comprising at least one nucleic acid ofclaim
 25. 56. A diagnostic kit for the detection of IAPs, preferablylivin-β, preferably in cancer cells, comprising a vector of claim 27.57. A diagnostic kit for the detection of IAPs, preferably livin-β,preferably in cancer cells, comprising a host cell of claim 28.